Imaging Spermine using LnDOTP5-Towards a Noninvasive Staging of Prostate Cancer
Abiola Olatunde1, Taylor Fuss1, Phillip Zhe Sun1, Leo L Cheng1, and Peter Caravan1

1Massachusetts General Hospital, Boston, MA, United States

Synopsis

Prostate cancer (PCa) is the most frequently diagnosed malignancy in men worldwide. Previous studies have indicated the utility of spermine as a potential biomarker for prostate cancer; however, quantifying spermine using MRS is difficult due to overlapping chemical shifts of spermine with other metabolites. We used LnDOTP5-, an anionic lanthanide macrocyclic complex, to form a stable ternary complex with positively-charged spermine to selectively shift spermine MR resonances. Here we report the affinity of different LnDOTP5- complexes for spermine and the effect of complex formation on spermine MR resonances in both D2O and serum solutions and intact human prostate tissue.

Target Audience

Prostate cancer (PCa) is the most frequently diagnosed malignancy in men worldwide, and the second leading cause of cancer death for men in the United States. Developing new PCa metabolomic biomarkers capable of predicting tumor aggressiveness can drastically improve PCa treatments and patient prognostications. Previous studies have indicated the utility of spermine as a potential biomarker for prostate cancer. However, quantifying spermine using in vivo MRS is difficult due to the overlapping chemical shifts of spermine with other metabolites in the spectra.

Purpose

We used LnDOTP5-, a stable anionic lanthanide macrocyclic complex, to form a sufficiently stable ternary complex with positively charged spermine. The ion-pair interaction resulted in the selective shift of the spermine MR resonances in D2O. The magnitude and direction of the shift is dependent on the pseudocontact contribution of the lanthanide (Ln). Here we report the affinity of different LnDOTP5- complexes (EuIII, YbIII,and TmIII) for spermine and the effect of complex formation on the spermine MR resonances in D2O and serum solutions, as well as intact human prostate tissue, to best mimic physiological conditions.

Methods

D2O and Serum Samples. Prepared samples of 10 mM spermine and 10 mM citrate in D2O or serum were analyzed with MRS on a Bruker AVANCE spectrometer operating at 600 MHz (14.1T). D2O was added to serum samples for field locking. EuIII, YbIII,and TmIII complexes were evaluated at 4ºC. Intact Tissue. Frozen tissue was scanned using high-resolution magic angle spinning (HRMAS) MRS on the same Bruker spectrometer. A 4 mm zirconia rotor with Kel-F inserts created a 10 μl sample space for tissue samples, and D2O was added for field locking. After an initial scan, 4 μl LnDOTP5- was added to one rotor and 4 μl of D2O was added to another rotor as control. Both rotors were kept overnight at 4ºC and then rescanned the next day. Spectra were recorded at 4ºC with the spectrometer frequency set on the water resonance. Spectra were measured with HRMAS with a spin rate of 3600Hz (±1.0Hz), and analyzed using an in-house developed MatLab based program.

Results

In D2O, spermine forms stable 1:1 complexes with LnDOTP5- (K > 105 M-1) and the lanthanide-induced shift can be large, with shifts up to 100 ppm for the TmIII complex at 37 ºC. In tissue, the spermine peaks were shifted, but the final locations were unknown (Figure 1). The resonances of citrate also disappeared, which was not seen in the control D2O solution. We further investigated these phenomena with solutions of bovine serum. In serum solutions, an excess of the LnDOTP5- is needed to achieve the maximum spermine shifts seen in a 1:1 ratio (LnDOTP5-: spermine) in D2O, suggesting other possible interactions with ions present within the serum, hence also probable in other physiological conditions (Figure 2). Further experiments, including the effects of cations and anions on the spermine shifts and the interactions between LnDOTP5- and citrate have also been conducted, with results to be presented at the meeting.

Discussion and Conclusions.

LnDOTP5- shift agent successfully shifted spermine MR resonances downfield of the water peak in both D2O and serum solutions. Spermine was also shifted away from its traditional resonance in tissue. The results of experiments exploring the effects of ions, such as Zn2+, Ca2+, and phosphate, as well as the observed shift of citrate in tissue, will enhance the understanding of the ion-pair interactions of spermine-LnDOTP5- complex in physiological conditions. The use of LnDOTP5- shift agents present as a possible noninvasive method for quantifying spermine and staging prostatic lesions, as well as a useful tool when combined with imaging techniques, such as CEST technology.

Acknowledgements

Authors acknowledge support by NIH grants CA115746 and EB009062, as well as the A. A. Martinos Center for Biomedical Imaging.

References

No reference found.

Figures

In intact human prostate tissue, YbDOTP5- shifted the spermine peaks to an unknown final location. The resonances of citrate also disappeared, which was not observed in the control tissue.

In bovine serum solutions, a 1:1 ratio of TmDOTP5-: spermine results in a significant downfield shift of spermine. Citrate is also slightly shifted, which was not observed in D2O solution, suggesting other possible interactions with ions present within the serum, hence also probable in other physiological conditions



Proc. Intl. Soc. Mag. Reson. Med. 24 (2016)
0470